DNA单分子近场光学成像与荧光探测

IMAGING AND FLUORESCENCE DETECTION OF DNA SINGLE MOLECULEUSING NEAR-FIELD OPTICAL MICROSCOPY

介绍了扫描近场光学(SNOM -Scanning Near-FieldOpticalMicroscope)/原子力显微镜(AFM -AtomicForceMicroscope)系统 (SNO/AM)的工作原理。在AFM模式和SNOM模式下对DNA分子进行成像和荧光探测 ,得到了清晰的DNA单分子的形貌像和荧光像。由形貌图像得到的DNA分子尺寸横向为20nm ,高度为2nm ,其中包含了探针形貌的影响。实验中采用Tapping模式的AFM成像 ,样品经多次搜索扫描无明显损坏。AFM模式的分辨率优于1nm。SNOM模式下DNA分子形貌像和荧光像清晰 ,由近场荧光分布可以确定分子取向和浓度。用YOYO -1染料对λDNA分子进行染色和荧光探测。通过对DNA分子多个截面进行测量 ,分析染料与DNA结合状态。

The operation principle and configuration of the Scanning Near-field Optical/Atomic Force Microscope (SNOM/AFM or SNO/AM) is described in this paper. DNA molecules were imaged and detected in the AFM mode and in the SNOM mode. The topography images and the fluorescence images of DNA single molecule have been obtained. The width of DNA molecule images is 20 nm and height 2 nm relevant to the shape of probe. No obvious damage of samples was observed under our experimental condition although several times of scanning in tapping mode were completed. The resolution of AFM is better than 1 nm. The topography images and the fluorescence images in SNOM mode are clear. The orientation and concentration of DNA molecules can be determined by the distribution of the near-field fluorescence images. λDNA molecules, in which YOYO-1 was intercalated, were imaged and characterized using the SNOAM. Areas of cross section in the DNA topography were measured. The intercalation and cooperation of YOYO-1 in the DNA were analyzed.